• Title/Summary/Keyword: cDNA 라이브러리

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Molecular Analysis of Alternative Transcripts of CCDC94 Gene in the Brain Tissues of Rhesus Monkey (붉은 털 원숭이의 뇌조직에서 CCDC94 유전자 대체 전사체의 분자적 분석)

  • Yun, Se-Eun;Ahn, Kung;Kim, Heui-Soo
    • Journal of Life Science
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    • v.21 no.3
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    • pp.459-463
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    • 2011
  • The genome of the rhesus monkey has diverged as an average sequence identity of ~93%. The rhesus monkey has been widely used as a non-human primate in the field of biomedical and evolutional research. Insertion of transposable elements (TEs) induced several events such as transcriptional diversity and different expression in host genes. In this study, 112 transcripts were identified from a full-length cDNA library of brain tissues of the rhesus monkey. One transcript (R54) showed a different expression pattern between human and rhesus monkey tissues. This phenomenon can be an explanation that R54 transcript was acquired by splicing a donor site derived from exonization of the L2A element. Therefore, integration of TEs during primate radiation could contribute to transcriptional diversity and gene regulation.

Functional Analysis of Expressed Sequence Tags from Hanwoo (Korean Cattle) cDNA Libraries (한우 cDNA 라이브러리에서 발현된 ESTs의 기능분석)

  • Lim, Da-Jeong;Byun, Mi-Jeong;Cho, Yong-Min;Yoon, Du-Hak;Lee, Seung-Hwan;Shin, Youn-Hee;Im, Seok-Ki
    • Journal of Animal Science and Technology
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    • v.51 no.1
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    • pp.1-8
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    • 2009
  • We generated 57,598 expressed sequence tags (ESTs) from 3 cDNA libraries of Hanwooo (Korean Cattle), fat, loin, liver. Liver, intermuscular fat and longissimus dorsi tissues were obtained from a 24-month-old Hanwoo steer immediately after slaughter. cDNA library was constructed according to the oligocapped method. The EST data were clustered and assembled into unique sequences, 4,759 contigs and 7,587 singletons. To carry out functional analysis, Gene Ontology annotation and identification of significant leaf nodes were performed that were detected by searching significant p-values from $2^{nd}$ level GO terms to leaf nodes using Bonferroni correction. We found that 13, 26 and 8 significant leaf nodes are unique in the transcripts according to 3 GO categories, molecular function, biological process and cellular component. Also digital gene expression profiling using the Audic's test was performed and tissue specific genes were detected in the above 3 libraries.

Analysis of Seed Hair Formation Related Genes by EST Profiling in Carrot (Daucus carota var. sativa) (EST profiling을 통한 당근(Daucus carota var. sativa)의 종모 형성에 관련된 유전자 분석)

  • Hwang, Eun-Mi;Oh, Gyu-Dong;Shim, Eun-Jo;Jeon, Sang-Jin;Park, Young-Doo
    • Horticultural Science & Technology
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    • v.28 no.6
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    • pp.1039-1050
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    • 2010
  • Carrot is one of the useful crops used abundantly in cooking in Western as well as Asia regions such as China and Korea. However, seed coats have hairs which should be removed to increase germination rate. Furthermore, because of seed hairs, farmers face several additional losses, such as time consumption, manpower, capital and so on, for seed handling. To prevent these problems, study of gene related hair formation using short-hair seed lines is required. We analyzed genes related to hair formation from seed through expressed sequenced tag (EST) profiling, based on the fact that the development of carrot seed hair is related to cellulose synthesis pathway in secondary cell wall synthesis stage. To study the gene expression related to hair formation of the carrot seed, a cDNA library was constructed by using the early maturation stage of the short-hair line (659-1) and hairy seed line (677-14). In short-hair (659-1) and hairy seed (677-14) lines, results from of EST profiling through BLASTX search analysis using the NCBI database showed that 172 and 224 unigenes had significant homology with known protein sequences, whereas 233 and 192 unigenes were not, respectively. All ESTs were grouped into 16 categories according to their putative functions. Twenty nine unigenes among all ESTs were considered to be genes regulating seed hair development from cellulose synthesis pathway during secondary cell wall synthesis stage; in results, 14 unigenes related to seed hair development were found only in hairy seed line.

Construction of Web-Based Database for Anisakis Research (고래회충 연구를 위한 웹기반 데이터베이스 구축)

  • Lee, Yong-Seok;Baek, Moon-Ki;Jo, Yong-Hun;Kang, Se-Won;Lee, Jae-Bong;Han, Yeon-Soo;Cha, Hee-Jae;Yu, Hak-Sun;Ock, Mee-Sun
    • Journal of Life Science
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    • v.20 no.3
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    • pp.411-415
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    • 2010
  • Anisakis simplex is one of the parasitic nematodes, and has a complex life cycle in crustaceans, fish, squid or whale. When people eat under-processed or raw fish, it causes anisakidosis and also plays a critical role in inducing serious allergic reactions in humans. However, no web-based database on A. simplex at the level of DNA or protein has been so far reported. In this context, we constructed a web-based database for Anisakis research. To build up the web-based database for Anisakis research, we proceeded with the following measures: First, sequences of order Ascaridida were downloaded and translated into the multifasta format which was stored as database for stand-alone BLAST. Second, all of the nucleotide and EST sequences were clustered and assembled. And EST sequences were translated into amino acid sequences for Nuclear Localization Signal prediction. In addition, we added the vector, E. coli, and repeat sequences into the database to confirm a potential contamination. The web-based database gave us several advantages. Only data that agrees with the nucleotide sequences directly related with the order Ascaridida can be found and retrieved when searching BLAST. It is also very convenient to confirm contamination when making the cDNA or genomic library from Anisakis. Furthermore, BLAST results on the Anisakis sequence information can be quickly accessed. Taken together, the Web-based database on A. simplex will be valuable in developing species specific PCR markers and in studying SNP in A. simplex-related researches in the future.

Construction of web-based Database for Haliotis SNP (웹기반 전복류 (Haliotis) SNP 데이터베이스 구축)

  • Jeong, Ji-Eun;Lee, Jae-Bong;Kang, Se-Won;Baek, Moon-Ki;Han, Yeon-Soo;Choi, Tae-Jin;Kang, Jung-Ha;Lee, Yong-Seok
    • The Korean Journal of Malacology
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    • v.26 no.2
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    • pp.185-188
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    • 2010
  • The Web-based the genus Haliotis SNP database was constructed on the basis of Intel Server Platform ZSS130 dual Xeon 3.2 GHz cpu and Linux-based (Cent OS) operating system. Haliotis related sequences (2,830 nucleotide sequences, 9,102 EST sequences) were downloaded through NCBI taxonomy browser. In order to eliminate vector sequences, we conducted vector masking step using cross match software with vector sequence database. In addition, poly-A tails were removed using Trimmest software from EMBOSS package. The processed sequences were clustered and assembled by TGICL package (TIGR tools) equipped with CAP3 software. A web-based interface (Haliotis SNP Database, http://www.haliotis.or.kr) was developed to enable optimal use of the clustered assemblies. The Clustering Res. menu shows the contig sequences from the clustering, the alignment results and sequences from each cluster. And also we can compare any sequences with Haliotis related sequences in BLAST menu. The search menu is equipped with its own search engine so that it is possible to search all of the information in the database using the name of a gene, accession number and/or species name. Taken together, the Web-based SNP database for Haliotis will be valuable to develop SNPs of Haliotis in the future.

Isolation and Characterization of a Nitric Oxide-induced Gene in Sweetpotato (고구마에서 질소 유도성 유전자의 분리 및 특성분석)

  • Lee, Il Hwan;Shim, Donghwan;Lee, Kang Lok;Nam, Ki Jung;Lee, Shin-Woo;Kim, Yun-Hee
    • Journal of Life Science
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    • v.29 no.6
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    • pp.631-636
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    • 2019
  • A new nitric oxide-induced (NOI) gene was isolated by screening ESTs from a cDNA library of dehydration-treated fibrous roots of sweetpotato (Ipomoea batatas). The 720 bp cDNA fragment, IbNOI, was sequenced, from which a 77 amino acid residue protein was deduced. A search of the protein BLAST database identified significant similarity to other plant NOI protein sequences. Quantitative RT-PCR analysis revealed diverse expression patterns of IbNOI in various tissues of the intact sweetpotato plant, and in leaves exposed to different stresses. The IbNOI gene was highly expressed in storage roots and suspension-cultured cells. In leaf tissues, IbNOI showed strong expression during sodium nitroprusside (SNP)-induced NO accumulation and chemical stress treatments. Expression of IbNOI was also induced under various abiotic stress conditions, such as dehydration, salt, and bacterial pathogen infection. These results suggest that IbNOI is involved in plant responses to diverse abiotic stresses and pathogen infection through a NO-related pathway.

Microbial Population Diversity of the Mud Flat in Suncheon Bay Based on 16S rDNA Sequences and Extracellular Enzyme Activities (남해안 갯벌 미생물의 세포외효소 활성 및 16S rDNA 분석에 의한 다양성 조사)

  • Kim, Yu-Jeong;Kim, Sung-Kyum;Kwon, Eun-Ju;Baik, Keun-Sik;Kim, Jung-Ho;Kim, Hoon
    • Applied Biological Chemistry
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    • v.50 no.4
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    • pp.268-275
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    • 2007
  • Diversity of the mud flat microbial population in Suncheon Bay was investigated by studying extracellular enzyme activities and 16S rDNA sequences. Four culturable bacterial strains with CMCase, xylanase and protease activities were isolated from the wetland and the mud flat. All the strains produced more xylanase activity than CMCase or protease activity, and the properties of the isolate enzymes from the wetland were similar to those from the mud flat. About 2,000 clones were obtained with the 16S rDNA amplified from the metagenomic DNA isolated from the mud samples. Based on the restriction pattern(s), seventeen clones were selected for base sequence analysis. Of the 17 clones, only 35% (6 clones) were found to be cultured strains and 65% (11 clones) to be uncultured strains. The similarities in the base sequences of the clones ranged from 91.0% to 99.9% with an average similarity of 97.3%. The clones could be divided into 7 groups, Proteobacteria (9 clones, 52.9%), Firmicutes (3 clones, 17.6%), Bacteroidetes (1 clone), Flavobacteria (1 clone), Verrucomicrobia (1 clone), Acidobacteria (1 clone), and Chloroflexi (1 clone). Most of the Proteobacteria clones were gamma Proteobacteria associated with oxidation-reduction of sulfur.

Purification and Gene Analysis of Peptidyl Prolyl cia-trans Isomerase from Bacillus stearothermophilus (Bacillus stearothermophilus Peptidyl Prolyl cis-trans Isomerase의 정제 및 유전자 분석)

  • 김동주
    • The Korean Journal of Food And Nutrition
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    • v.15 no.2
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    • pp.104-111
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    • 2002
  • The peptidyl prolyl sis-trans isomerase (PPIase, EC 5.2.1.8) from bacillus stearothermophilus was extracted from the cells treated with by lysozyme. PPIase was purified from the cell extracts by heat treatment, ammonium sulfate precipitation, ion exchange chromatography and finally gel filtration, sodium dodecyl sulfate polyacrylamide gel electrophoresis(SDS-PAGE). The molecular weight of the purified PPIase was estimated as 18kDa by SDS-PAGE. The 39 amino acid residues from the N-terminus were determined by the protein sequencer. The enzyme showed the optimum pH at 8.0 and was stable at the range of pH 7.0∼8.0. The enzyme was considerably stable after heat treatment at 60$\^{C}$ for 30minutes, and the enzyme was quite stable up to 65$\^{C}$. The presence of the PPIase in the refolding solution accelerated the isomerization rate of the assay peptide. PPIase gene of Bacillus stearothermophilus was screened from a genomic library by plaque hybridization using the A-l primer as a probe. A PPIase positive plaque contained a 3.0kb insert of the chromosomal DNA. A 3.0kb fragment was subcloned into pUC18, resulting pPI-40. A DNA fragment encoding the N-terminal portion of the PPIase in pPI-40 was amplified by polymerase chain reaction(PCR) method using the A-1 and B-2 primers. The amplified fragment was cloned into the Sma I site of pUC18 and recombinant plasmid was designated as pSN-18. The nucleotide sequence of 167bp fragment was determined. The deduced amino acid sequence of PPIase was completely matched with the determined N-terminal amino acid sequence of PPIase B. stearothermophilus.

Construction of Genetic Linkage Map and Identification of Quantitative Trait Loci in Populus davidiana using Genotyping-by-sequencing (Genotyping-by-sequencing 기법을 이용한 사시나무(Populus davidiana) 유전연관지도 작성 및 양적형질 유전자좌 탐색)

  • Suvi Kim;Yang-gil Kim;Dayoung Lee;Hye-jin Lee;Kyu-Suk Kang
    • Journal of Korean Society of Forest Science
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    • v.112 no.1
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    • pp.40-56
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    • 2023
  • Tree species within the Populus genus grow rapidly and have an excellent capacity to absorb carbon, conferring substantial ability to effective purify the environment. Poplar breeding can be achieved rapidly and efficiently if a genetic linkage map is constructed and quantitative trait loci (QTLs) are identified. Here, a high-density genetic linkage map was constructed for the control pollinated progeny using the genotyping-by-sequencing (GBS) technique, which is a next-generation sequencing method. A search was also performed for the genes associated with quantitative traits located in the genetic linkage map by examining the variables of height and diameter at root collar, and resilience to insect damage. The height and diameter at root collar were measured directly, while the ability to recover from insect damage was scored in a 4-year-old breeding population of aspen hybrids (Odae19 × Bonghyeon4 F1) established in the research forest of Seoul National University. After DNA extraction, paternity was confirmed using five microsatellite markers, and only the individuals for which paternity was confirmed were used for the analysis. The DNA was cut using restriction enzymes and the obtained DNA fragments were prepared using a GBS library and sequenced. The analyzed results were sorted using Populus trichocarpa as a reference genome. Overall, 58,040 aligned single-nucleotide polymorphism (SNP) markers were identified, 17,755 of which were used for mapping genetic linkages. The genetic linkage map was divided into 19 linkage groups, with a total length of 2,129.54 cM. The analysis failed to identify any growth-related QTLs, but a gene assumed to be related to recovery from insect damage was identified on linkage group (chromosome) 4 through genome-wide association study.

EST Profiling for Seed-hair Characteristic and Development of EST-SSR and SNP Markers in Carrot (당근 종모 형질 관련 EST profiling과 이를 이용한 EST-SSR 및 SNP 마커 개발)

  • Oh, Gyu-Dong;Hwang, Eun-Mi;Shim, Eun-Jo;Jeon, Sang-Jin;Park, Young-Doo
    • Horticultural Science & Technology
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    • v.28 no.6
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    • pp.1025-1038
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    • 2010
  • Carrot ($Daucus$ $carota$ L. var. $sativa$) is one of the most widely used crops in the world. Moreover it is an important crop because of its high content of ${\beta}$-carotene, well-known as the precursor of vitamin A carotenoid. However, seed-hair which is generated in epidermal cell of seeds inhibits absorption and germination. For that reason, carrot seeds are commercialized after mechanical hair removal process. To overcome such cumbersome weaknesses, new breeding program for developing hairless-seed carrot cultivar has been needed. Therefore, in this study, cDNA libraries from seeds of short-hair seed phenotype CT-ATR615 OP 666-13line and hairy seed CT-ATR615 OP-CK1-9 line were constructed and expression patterns related to generation of seed-hair were analyzed by comparison of EST sequences. Differential EST sequence results between two lines were classified into FunCat functional categories based on the results of BlastX search. Higher expression quantities belonging to metabolic category were shown on short-hair seed line than hairy-seed one. Differential expression quantities between those two lines in the protein folding and stabilization, subcellular localization categories were supposed to contribute variously on the generation of seed-hair. We confirmed 50 and 59 SSR sites, and 2 SNP sites by analyzing EST sequences in two lines; thereafter, we designed SNP and SSR primer sets from these EST sequence information as a molecular marker. These markers are thought to be used in research of molecular markers for classification of carrot family and related to various traits, as well as seed-hair characteristic.